Global and Complement Gene-Specific DNA Methylation in Grass Carp after Grass Carp Reovirus (GCRV) Infection
Abstract
:1. Introduction
2. Results
2.1. Grass Carp after GCRV Infection
2.2. Global DNA Methylation Changes after GCRV Infection
2.3. Expression Patterns of DNA Methyltransferase Genes after GCRV Infection
2.4. Expression Patterns of Ten-Eleven Translocation Proteins and Glycine N-methyltransferase after GCRV Exposure
2.5. Expression Patterns of Representative Complement Genes after GCRV Infection
2.6. Methylation Changes in the 5′ flanking Region of C3 and KNG1 after GCRV Infection
2.7. The Relationship between Expression Level and DNA Methylation Level of C3 and KNG1 after GCRV Infection
3. Discussion
4. Materials and Methods
4.1. Ethical Procedures
4.2. Experimental Animals, Virus Exposure, and Sample Collection
4.3. DNA Isolation and Global DNA Methylation Measurement
4.4. Bisulfite Treatment and Sequencing
4.5. RNA Isolation and Quantitative Real-Time PCR (RT-qPCR)
4.6. Statistical Analysis
Acknowledgments
Author Contributions
Conflicts of Interest
Abbreviations
C3 | complement component 3 |
CpG | cytosine-phosphor-guanine |
DNMT | DNA methyltransferase |
GCRV | grass carp reovirus |
GNMT | glycine N-methyltransferase |
KNG1 | kininogen-1 |
LSD | least significant difference |
MDV | Marek’s disease virus |
TETs | ten-eleven translocation proteins |
TIS | translation initiation site |
TSS | transcriptional start site |
SAM | S-adenosyl-l-methionine |
SPSS | Statistical Product and Service Solutions |
5hmC | 5-hydroxymethylcytosine |
5-mC | 5-methylcytosine |
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Primers | Sequences (5′ to 3′) | Usage |
---|---|---|
GCRV-F | GGCGGCTGCTATGATACCAGTAGAC | GCRV detection |
GCRV-R | GTCGCTTCAGATCAATCACCAAATC | |
C3-BSP-F | TTAGAGGGTTATTATGTTGAGTATTT | BSP PCR of C3 |
C3-BSP-R | AAAACTCATAAAACCATTCAACTC | |
C3-BSP-outer-R | AACAATATAAATTTTTCAAAAAATTC | |
KNG1-BSP-F | TGTAAAGGTTTTTGTGAAGGTTTTT | BSP PCR of KNG1 |
KNG1-BSP-R | CCACTAAACTCTATCCTCCCCTATAATA | |
KNG1-BSP-outer-R | TCTCTACTCTTCTAAAAACCCAACAA | |
Qdnmt1-F | CCACCGAAATGTGCCGACTG | QPCR of dnmt1 |
Qdnmt1-R | GGTGCCCATGCTTGTCATACAC | |
Qdnmt2-F | AAGTCTCAAAGCCCTTCACATTCTC | QPCR of dnmt2 |
Qdnmt2-R | ACTACCCCACTCCTGCCAAAAAC | |
Qdnmt6-F | CAACCGCCACAGCCTCAAACT | QPCR of dnmt6 |
Qdnmt6-R | CCTCGGCCATCTCGTACTCTGAC | |
Qdnmt7-F | GAGAAGCAAGTGGGCTCCTACG | QPCR of dnmt7 |
Qdnmt7-R | CAGCGATGGAGGTCGTAAGAGTT | |
Qtet1-F | AGGCGTGTGAACATCAGGTGG | QPCR of tet1 |
Qtet1-R | AACCGAAAAACGGGGCATC | |
Qtet2-F | CCTAACTCCAAAATAGACGGCAC | QPCR of tet2 |
Qtet2-R | GATGCGGGGTCATTGGTTTA | |
Qtet3-F | GCCGCATATCCCTGGTCTTCTA | QPCR of tet3 |
Qtet3-R | TCGTGTCATCACACCGTCTCGT | |
Qgnmt-F | AGTCTGAATGCCCTCTGGTGG | QPCR of gnmt |
Qgnmt-R | ACCAGACTTCAAAGGGACCAAA | |
QC3-F | ATGGTTCGCAAACACTCCTCAG | QPCR of C3 |
QC3-R | ACAGGTACTTGGCTTCTATGTCAACT | |
Qkng1-F | TGAAGGTGCTCAACTGGCTCT | QPCR of kng1 |
Qkng1-R | AGTAACTCTTGACCTGCTTCTGTAAAC | |
Q-β-actin-F | AGCCATCCTTCTTGGGTATG | QPCR of β-actin |
Q-β-actin-R | GGTGGGGCGATGATCTTGAT | |
Q-GCRV-F | AGCGCAGCAGGCAATTACTATCT | QPCR of GCRV |
Q-GCRV-R | ATCTGCTGGTAATGCGGAACG |
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Xiong, L.; He, L.; Luo, L.; Li, Y.; Liao, L.; Huang, R.; Zhu, Z.; Wang, Y. Global and Complement Gene-Specific DNA Methylation in Grass Carp after Grass Carp Reovirus (GCRV) Infection. Int. J. Mol. Sci. 2018, 19, 1110. https://doi.org/10.3390/ijms19041110
Xiong L, He L, Luo L, Li Y, Liao L, Huang R, Zhu Z, Wang Y. Global and Complement Gene-Specific DNA Methylation in Grass Carp after Grass Carp Reovirus (GCRV) Infection. International Journal of Molecular Sciences. 2018; 19(4):1110. https://doi.org/10.3390/ijms19041110
Chicago/Turabian StyleXiong, Lv, Libo He, Lifei Luo, Yongming Li, Lanjie Liao, Rong Huang, Zuoyan Zhu, and Yaping Wang. 2018. "Global and Complement Gene-Specific DNA Methylation in Grass Carp after Grass Carp Reovirus (GCRV) Infection" International Journal of Molecular Sciences 19, no. 4: 1110. https://doi.org/10.3390/ijms19041110